Advanced glycation endproduct modified basement membrane attenuates endothelin-1 induced [Ca2+]i signalling and contraction in retinal microvascular pericytes

Sarah Jane Hughes, N. Wall, C. N. Scholfield, Graham McGeown, T. A. Gardiner, A. W. Stitt, T. M. Curtis

Research output: Contribution to journalArticle

18 Citations (Scopus)

Abstract

PURPOSE: To assess the effects of advanced glycation endproduct (AGE) modification of vascular basement membrane (BM) on endothelin-1 (Et-1) induced intracellular [Ca2+] ([Ca2+]i) homeostasis and contraction in retinal microvascular pericytes (RMP). METHODS: RMPs were isolated from bovine retinal capillaries and propagated on AGE modified BM extract (AGE-BM) or non-modified native BM. Cytosolic Ca2+ was estimated using fura-2 microfluorimetry and cellular contraction determined by measurement of planimetric cell surface area. ETA receptor mRNA and protein expression was assessed by real time RT-PCR and western blotting, respectively. RESULTS: Exogenous endothelin-1 (Et-1) evoked rises in [Ca2+]i and contraction in RMPs were found to be mediated entirely through ETA receptor (ETAR) activation. Both peak and plateau phases of the Et-1 induced [Ca2+]i response and contraction were impaired in RMPs propagated on AGE modified BM. ETAR mRNA expression remained unchanged in RMPs exposed to native or AGE-BM, but protein expression for ETAR (66 kDa) was lower in the AGE exposed cells. CONCLUSIONS: These results suggest that substrate derived AGE crosslinks can influence RMP physiology by mechanisms which include disruption of ETA receptor signalling. AGE modification of vascular BMs may contribute to the retinal hemodynamic abnormalities observed during diabetes.
Original languageUndefined/Unknown
Pages (from-to)996-1004
Number of pages9
JournalMolecular Vision
Volume10
Publication statusPublished - 27 Dec 2004

Bibliographical note

LR: 20061115; JID: 9605351; 0 (Endothelin-1); 0 (Glycosylation End Products, Advanced); 0 (RNA, Messenger); 0 (Receptor, Endothelin A); 7440-70-2 (Calcium); 96314-98-6 (Fura-2); epublish

Keywords

  • Animals
  • Basement Membrane/drug effects/metabolism
  • Blotting, Western
  • Calcium/metabolism
  • Calcium Signaling/drug effects
  • Capillaries
  • Cattle
  • Contracture/metabolism
  • Electrophoresis, Polyacrylamide Gel
  • Endothelin-1/pharmacology
  • Fura-2/metabolism
  • Glycosylation End Products, Advanced/pharmacology
  • Microscopy, Fluorescence
  • Pericytes/cytology/physiology
  • RNA, Messenger/metabolism
  • Receptor, Endothelin A/genetics
  • Retinal Vessels/cytology/physiology
  • Reverse Transcriptase Polymerase Chain Reaction

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